Therapeutic agent for inflammatory bowel disease and tnf-alpha production inhibitor

ABSTRACT

Disclosed is an agent for use in the treatment or prevention of inflammatory bowel disease. Also disclosed is an agent for inhibiting the production of TNF-α. A therapeutic or prophylactic agent for inflammatory bowel disease comprising at least one amino acid selected from the group consisting of lysine, histidine, phenylalanine, methionine, tryptophan, glutamine, glycine, cysteine, cystine and threonine, the amino acid being administered at a dose of 0.1 to 4000 mg/kg per day; and a TNF-α production inhibitor comprising an amino acid selected from the group consisting of histidine, phenylalanine and tryptophan, the amino acid being administered at a dose of 0.1 to 4000 mg/kg per day.

TECHNICAL FIELD

The present invention relates to an agent for treating or preventing aninflammatory bowel disease, comprising a particular amino acid(s). Thepresent invention also relates to an agent for inhibiting TNFproduction, comprising a particular amino acid(s).

BACKGROUND ART

An inflammatory bowel disease is a generic term for enteropathy withinflammation, and mainly includes ulcerative colitis and Crohn'sdisease. The ulcerative colitis is a diffuse non-specific inflammatorydisease where large bowel mucosa or submucosa is affected and erosion orulcer is often formed. Clinical symptoms are mucous and bloody stools,abdominal pain, blood stools, watery stools, fever, lack of appetite,malevolence, emesis and the like. As agents for treating the ulcerativecolitis, salazosulfapyridine, adrenal cortex steroids,immunosuppressants, 5-aminosalicylic acid (5-ASA) and the like are used,but it can not be said that these agents are enough to treat theulcerative colitis.

Crohn's disease is an idiopathic chronic enteritis of unknown cause, andexhibits non-specific inflammatory symptoms in intestines from a smallintestine to a large intestine. Its lesions are composed ofgranulomatous lesions with fibrosis and ulcer, and it is likely that thelesions appear in all gastrointestinal area from an oral cavity to ananus. The clinical symptoms of Crohn's disease include abdominal pain,general malaise, diarrhea, melena, occult blood positive, fever, weightloss, anemia, ileus symptom, abdominal tumor, malevolence, emesis andperitonitis symptom. Crohn's disease simultaneously causes variousgastrointestinal and parenteral symptoms, e.g., intestinal stenosis,intestinal perforation, abdominal abscess and heavy hemorrhage which areserious conditions, in addition to nutritional disorder, and oftenrequires procedures such as intestinal surgery. In Japan, a high calorieinfusion or an enteral nutrition is performed for the purpose ofimproving the nutritional condition. In the high calorie infusion, arisk of bacterial translocation is increased. Thus, particularly for along term, the enteral nutrition is performed. In addition, the therapyby an agent has been attempted. In the drug therapy,salazosulfapyridine, metronodazole, adrenal cortex steroids,immunosuppressants, 5-aminosalicylic acid (5-ASA) and the like areadministered. Recently, an anti-TNF antibody has begun to beadministered clinically. However, it can be said that the administrationof these agents is insufficient yet for treating the Crohn's disease.

TNF-α is an inflammatory cytokine produced by macrophages, macrophagelineage cells (Kupper cells), neutrophils, basophils, eosinophils,lymphocytes, NK cells, LAK cells, mast cells, bone marrow cells,fibroblasts, astrocytes, keratinocytes and the like, and has beenrecently demonstrated to be deeply involved in pathogenesis of manydiseases including Crohn's disease. Therefore, it is believed that ifthe action of TNF-α can be inhibited, it becomes possible to treat thosediseases. Currently, steroidal hormone agents and non-steroidalanti-inflammatory agents are applied to some inflammatory diseases.However, they have diverse action points and do not have an inhibitoryaction specific for TNF-α. Thus, it is likely to elicit harmful sideeffects. Particularly, the side effect of the steroid agent has become amedical problem. Furthermore, the treatment using an anti-TNF-α antibodyand a soluble TNF-α receptor which are peptide macromolecules gives goodclinical results in chronic rheumatoid arthritis and Crohn's disease,but sometimes induces serious infectious diseases including tuberculosisand sepsis, and deterioration of demyelinating disease. Occurrence ofmalignant tumors has been also reported. Additionally, the formation ofa neutralization antibody has been reported, and thus they can not besaid to be sufficient.

DISCLOSURE OF INVENTION Problem to be Solved by the Invention

The present invention aims at providing an agent for use in thetreatment or prevention of inflammatory bowel diseases. The presentinvention also aims at providing an agent for inhibiting the productionof TNF-α.

Means for Solving Problem

As a result of an extensive study on inflammatory bowel diseases andTNF-α production for accomplishing the above objects, the presentinventor has found that an excellent effect is obtained by administeringa particular amino acid(s) in a particular amount(s), and completed thepresent invention.

That is, the present invention provides an agent for treatment orprevention of an inflammatory bowel disease wherein it contains one ormore amino acids selected from the group consisting of lysine,histidine, phenylalanine, methionine, tryptophan, glutamine, glycine,cysteine, cystine and threonine, and the amino acids are administered inan amount of 0.1 to 4000 mg/kg per day.

The individual amino acid described herein includes any form of D-type,L-type or a mixture of D and L, and preferably the L-type is used. Theindividual amino acid may also be in a salt form in addition to the freeamino acid. Furthermore, the individual amino acid may be the form of apeptide. In the case of the peptide, the number of the amino acids ispreferably 20 or less. A dosage of the amino acid is given in terms ofthe free amino acid.

The present invention provides an agent for treatment or prevention ofan inflammatory bowel disease wherein it contains 0.5 to 2% by weight oftryptophan, 3 to 9% by weight of lysine, 1.5 to 5% by weight ofhistidine, 4 to 10% by weight of phenylalanine and 9 to 23% by weight ofglutamine, and the amino acids are administered in an amount of 0.1 to4000 mg/kg per day.

Furthermore, the present invention provides an inhibitor of TNF-αproduction wherein it contains amino acids selected from the groupconsisting of histidine, phenylalanine and tryptophan, and the aminoacids are administered in an amount of 0.1 to 4000 mg/kg per day.

Still further, the present invention provides an inhibitor of TNF-αproduction wherein it contains 0.5 to 2% by weight of tryptophan, 3 to9% by weight of lysine, 1.5 to 5% by weight of histidine, 4 to 10% byweight of phenylalanine and 9 to 23% by weight of glutamine, and theamino acids are administered in an amount of 0.1 to 4000 mg/kg per day.

BEST MODES FOR CARRYING OUT THE INVENTION

The inflammatory bowel disease to which the agent for treatment orprevention of the present invention is applied is an intestine-relateddisease, and the agent is effective for example for ulcerative colitisand Crohn's disease.

The agent for treatment or prevention of the inflammatory bowel diseaseof the present invention contains the amino acids selected from thegroup consisting of lysine, histidine, phenylalanine, methionine,tryptophan, glutamine, glycine, cysteine, cystine and threonine. One ormore amino acids may be contained. In light of therapeutic or preventiveeffect, the agent for treatment or prevention of the inflammatory boweldisease of the present invention preferably contains the amino acidsselected from the group consisting of lysine, histidine, phenylalanine,tryptophan and glutamine, and most preferably contains tryptophan. Oneor more amino acids selected from the group consisting of lysine,histidine, phenylalanine, and glutamine may be contained in addition totryptophan.

The agent for treatment or prevention of inflammatory bowel disease ofthe present invention is administered in the amount of 0.1 to 4000 mg/kgof the amino acid(s) (when two or more amino acids are contained, theamount is a total thereof) per day. In light of therapeutic orpreventive effect, the agent for treatment or prevention of theinflammatory bowel disease of the present invention is preferablyadministered in the amount of 0.1 to 1000 mg/kg and most preferably 1 to500 mg/kg of the amino acid(s) per day.

Furthermore, the amino acids other than the above, e.g., isoleucine,leucine, valine, arginine, alanine, aspartic acid, proline, serine,tyrosine, glutamic acid, asparagine and the like may be contained.

In another embodiment, the agent for treatment or prevention of theinflammatory bowel disease of the present invention contains 0.5 to 2%by weight of tryptophan, 3 to 9% by weight of lysine, 1.5 to 5% byweight of histidine, 4 to 10% by weight of phenylalanine, and 9 to 23%by weight of glutamine. In light of therapeutic or preventive effect,the agent for treatment or prevention of the inflammatory bowel diseaseof the present invention contains 0.9 to 1.5% by weight of tryptophan, 4to 7.1% by weight of lysine, 2 to 4% by weight of histidine, 5 to 9% byweight of phenylalanine, and 11 to 20% by weight of glutamine, and mostpreferably contains 1.0 to 1.3% by weight of tryptophan, 4.5 to 6% byweight of lysine, 2.4 to 3.5% by weight of histidine, 6 to 8% by weightof phenylalanine, and 13 to 17% by weight of glutamine.

The agent for treatment or prevention of inflammatory bowel disease isadministered in the amount of 0.1 to 4000 mg/kg per day in terms of thetotal amount of the amino acids. In light of therapeutic or preventiveeffect, the agent for treatment or prevention of the inflammatory boweldisease is administered in the amount of 0.1 to 1000 mg/kg and mostpreferably 1 to 500 mg/kg per day in terms of the total amount of theamino acids.

Furthermore, the amino acids other than the above, e.g., isoleucine,leucine, methionine, threonine, valine, arginine, alanine, asparticacid, glycine, proline, serine, tyrosine, cysteine, cystine, glutamicacid, asparagine and the like may be contained.

The inhibitor of the present invention inhibits the secretion of TNF-αfrom TNF-α producing cells such as macrophages, macrophage lineage cells(Kupper cells), neutrophils, basophils, eosinophils, lymphocytes, NKcells, LAK cells, mast cells, bone marrow cells, fibroblasts,astrocytes, keratinocytes and the like. Therefore the inhibitor of thepresent invention is anticipated as being usable as the agent fortreatment or prevention of the diseases where the inhibition of TNF-αproduction is effective, e.g., sepsis, septic shock, endotoxin shock,oligemic shock, post-oligemic reperfusion injury, meningitis, psoriasis,congestive heart failure, fibrosis, hepatitis, insulin independentdiabetes, graft rejection, graft versus host disease, cancer, cachexia,arthritis (chronic rheumatoid arthritis, rheumatoid spondylitis,osteoarthritis, other arthritis), inflammatory bone diseases, boneresorption diseases, Behcet's syndrome, infectious diseases(opportunistic infection due to AIDS, cerebral malaria, infection withMycobacterium), autoimmune diseases (systemic lupus erythematosus,rheumatoid diseases, allergy, multiple sclerosis, autoimmune uveitis,nephrosis syndrome, type I diabetes (IDDM)), Crohn's disease, ulcerativecolitis, erythema nodosum, and damage of alveoli due to radiationdisorder and hyperoxia.

The inhibitor of the TNF-α production of the present invention containsthe amino acids selected from the group consisting of histidine,phenylalanine and tryptophan. One or more of the amino acids may becontained.

The inhibitor of the TNF-α production is administered in the amount of0.1 to 4000 mg/kg of the amino acid(s) (when two or more amino acids arecontained, the amount is the total thereof) per day. In light ofinhibitory effect, the inhibitor of the TNF-α production is preferablyadministered in the amount of 0.1 to 1000 mg/kg and most preferably 1 to500 mg/kg of the amino acid(s) per day.

Furthermore, the amino acids other than the above, e.g., lysine,methionine, glutamine, glycine, cysteine, cystine, threonine,isoleucine, leucine, valine, arginine, alanine, aspartic acid, proline,serine, tyrosine, asparagine, glutamic acid and the like may becontained.

In another embodiment, the inhibitor of the TNF-α production of thepresent invention contains 0.5 to 2% by weight of tryptophan, 3 to 9% byweight of lysine, 1.5 to 5% by weight of histidine, 4 to 10% by weightof phenylalanine and 9 to 23% by weight of glutamine. In light ofinhibitory effect, the inhibitor of the TNF-α production of the presentinvention preferably contains 0.9 to 1.5% by weight of tryptophan, 4 to7.1% by weight of lysine, 2 to 4% by weight of histidine, 5 to 9% byweight of phenylalanine and 11 to 20% by weight of glutamine, and mostpreferably contains 1.0 to 1.3% by weight of tryptophan, 4.5 to 6% byweight of lysine, 2.4 to 3.5% by weight of histidine, 6 to 8% by weightof phenylalanine and 13 to 17% by weight of glutamine.

The inhibitor of the TNF-α production is administered in the amount of0.1 to 4000 mg/kg per day in terms of the total amount of the aminoacids. In light of inhibitory effect, the inhibitor of the TNF-αproduction is preferably administered in the amount of 0.1 to 1000 mg/kgand most preferably 1 to 500 mg/kg per day in terms of the total amountof the amino acids.

Furthermore, the amino acids other than the above, e.g., methionine,glycine, cysteine, cystine, threonine, isoleucine, leucine, valine,arginine, alanine, aspartic acid, proline, serine, tyrosine, asparagine,glutamic acid ant the like may be contained.

The agent for treatment or prevention of the inflammatory bowel diseaseand the inhibitor of the TNF-α production of the present invention maybe appropriate formulations, and are prepared in the form of, forexample, powders, particles, granules, tablets, capsules and liquids.

As additives added to the powders, particles, granules, tablets andcapsules, for example, excipients (e.g., lactose, glucose, D-mannitol,starch, crystalline cellulose, calcium carbonate, kaolin, light silicacid anhydrate, trehalose), binders (e.g., starch glue liquid, gelatinsolution, hydroxypropylcellulose, hydroxypropylmethylcellulose,polyvinyl pyrrolidone, ethanol), disintegrants (e.g., starch, gelatinpowder, carboxymethylcellulose, carboxymethylcellulose calcium salt),lubricants (e.g., magnesium stearate, talc), coating agents (e.g.,hydroxypropylcellulose, hydroxypropylmethylcellulose, acetylcellulose,saccharose, titanium oxide) are available. Additionally if necessary,coloring agents, flavoring agents and odor improving agents are added.As the additives added to oral liquid agents, preservatives (e.g.,benzoic acid, paraoxybenzoate ester, sodium dehydroacetate), suspendingagents and emulsifiers (e.g., gum arabic, tragacanth,carboxymethylcellulose sodium salt, methylcellulose, egg yolk,surfactant), and sweeteners and acidifiers (e.g., trehalose, citricacid) are available. Additionally, if necessary, coloring agents andstabilizers are added. As solvents used therefor, purified water ismainly used, but ethanol, glycerine and propylene glycol can also beused.

Additionally, nutritional ingredients such as dextrin, protein sources,carbohydrate sources, vitamins, minerals and trace elements may becontained. The protein source may be the protein useful for nutritionsupply, and may be any of the animal proteins and plant proteins. As theanimal protein, milk proteins are preferable, and particularly lowlactose milk proteins and casein are preferable. As the plant protein, aseparated soybean protein is preferable. Two or more proteins may becombined. The protein source may be peptides obtained by hydrolyzing theprotein. As the carbohydrate source, saccharides are preferable,monosaccharides, disaccharides, and polysaccharides can be included, andmore specifically, glucose, fructose, mannose, galactose, sucrose, sugar(may be purified saccharose), maltose, lactose, dextrin, maltodextrin,starch, maize starch, soybean oligosaccharide and sugar alcohol can beincluded. Two or more of these saccharides may be combined. As thecarbohydrate source, it is preferable to contain any one of sugar,dextrin, maltodextrin and maize starch. The vitamins include vitamin B1,vitamin B2, vitamin B6, vitamin B12, vitamin C, vitamin A, vitamin D,vitamin E, vitamin K, vitamin H, folic acid, pantothenic acids andnicotinic acids. Particularly, vitamin C and vitamin E are preferablebecause they have a property as an antioxidant. The minerals are notparticularly limited as long as they are generally used in this field.Specifically, calcium, sodium, potassium, magnesium, chlorine andphosphorus in an inorganic or organic salt form can be included. Foreach inorganic or organic salt, the same salts as those which have beenalready placed on the market and combined in the infusion and theenteral nutrition can be used. The trace element is a metal elementwhich is a trace amount but indispensable for a living body.Specifically, zinc, iron, manganese, copper, chromium, molybdenum,serene, fluorine and iodine in the inorganic and organic salt form areincluded. Each trace element may be combined in consideration of a dailyneeded amount.

The agent for treatment or prevention of the inflammatory bowel diseaseand the inhibitor of the TNF-α production of the present invention canbe prepared by standard methods of granulating each active ingredientdirectly or mixing each active ingredient with the pharmaceuticallyacceptable additives depending on each formulation and granulating, ordissolving the agent in the appropriate solvent to emulsify or suspendit, and further mixing it with an appropriate base.

The agent for treatment or prevention of the inflammatory bowel diseaseand the inhibitor of the TNF-α production of the present invention canbe administered orally or parenterally. Contents of the above aminoacids, additives and nutritional ingredients are controlled so that theaforementioned amount to be administered daily can be accomplished bydosing once or several times daily. The preferable method ofadministration includes oral administration, enteral administration(trans-gastric administration, trans-duodenal administration,percutaneous endoscopic gastrostomy (PEG) using a tube, or an enema ispreferable), and intravenous administration.

EXAMPLES Therapeutic Effect on Inflammatory Bowel Disease

Balb/c background IL-10 deficient mice were established from C57BL/6background IL-10 deficient mice by back-crossing for seventhgenerations. Balb/c background IL-10 deficient mice spontaneouslydevelop the colitis with aging and exhibit symptoms such as diarrhea.Spleen, mesenteric lymph nodes and sacral lymph nodes were collectedfrom the IL-10 gene-deficient Balb/c mice (male) which had developed thecolitis detected by observing the diarrhea symptoms. After preparingsingle cell suspensions, the cells at about 1×10⁷ were injectedintraperitoneally to Scid mice which were immunodeficiency mice(hereinafter the Scid mouse injected IP was referred to as an“experiment mouse”). Subsequently, a chow was changed to an experimentaldiet. As the experimental diet, a standard chow or those obtained bymixing the amino acids with the standard chow shown in the followingTables 1 to 5 were used. At a time point three weeks after celltransfer, the mice were killed, the colon was collected, the intestinalcontents were washed out, and its weight was measured. A percentage ofinhibiting a weight gain of the colon was calculated by the followingformula.

Percentage of inhibiting a weight gain of colon=[1−(Colon weight intarget group−Colon weight in cell transfer group)/(Colon weight instandard chow group−Colon weight in cell transfer group)]×100(%)

Results are shown in the following Tables 1 to 5. An amino acid mixtureA is composed of the following amino acids (% by weight is representedin terms of free amino acid).

L-isoleucine: 4.56% by weightL-leucine: 6.38% by weightL-lysine hydrochloride: 6.30% by weightL-methionine: 4.60% by weightL-phenylalanine: 6.18% by weightL-threonine: 3.71% by weightL-tryptophan: 1.07% by weightL-valine: 4.98% by weightL-histidine hydrochloride: 3.56% by weightL-arginine hydrochloride: 7.99% by weightL-alanine: 6.38% by weightMagnesium potassium L-aspartate: 7.35% by weightSodium aspartate hydrate: 6.15% by weightL-glutamine: 13.71% by weightGlycine: 3.58% by weightL-proline: 4.47% by weightL-serine: 8.23% by weight andL-tyrosine: 0.78% by weight

TABLE 1 Normal Control Example 1 Non- Standard Standard chow + treatmentchow Gln (5%) Colon Mean 248.6 612.5 530.5 weight SE 5.2 18.2 14.6 n 8 88 Rate of 22.5 inhibiting weight gain of colon (%)

TABLE 2 Normal Control Example 2 Example 3 Example 4 Non- Stan- StandardStandard Standard treat- dard chow + chow + chow + ment chow Lys(5%)His(5%) Cys(5%) Colon Mean 219.2 556.4 392.8 429.0 414.4 weight SE 8.369.3 30.8 41.1 27.4 n 7 7 7 7 7 Rate of 48.5 37.8 42.1 inhibiting weightgain of colon (%)

TABLE 3 Example 5 Example 6 Example 7 Example 8 Normal Control StandardStandard Standard Standard non- Standard chow + chow + chow + chow +treatment chow Gly(5%) Trp(5%) Met(5%) Cys(5%) Colon weight Mean 210.1573.0 437.1 236.7 307.7 375.4 SE 6.3 29.0 32.0 22.9 25.3 12.8 n 7 7 7 77 7 Rate of 37.4 92.7 73.1 54.4 inhibiting weight gain of colon (%)

TABLE 4 Example Example Example 9 10 11 Standard standard StandardExample Example chow + chow + chow + 12 13 Normal Control Amino acidAmino acid Amino acid Standard Standard Non- Standard mixture A mixtureA mixture A chow + chow + treatment chow (10%) (20%) (30%) Phe(5%)Thr(5%) Colon weight Mean 219.2 556.4 488.6 455.5 383.3 392.8 468.2 SE8.3 69.3 16.2 22.6 29.2 30.8 13.2 n 7 7 7 7 7 7 7 Rate of 23.4 32.1 51.148.5 28.8 inhibiting weight gain of colon (%)

TABLE 5 Example 14 Example 15 Normal Control Standard Standard Non-Standard chow + chow + treatment chow Trp (2%) Trp (5%) Colon Mean 204.8585.0 425.6 262.1 weight SE 6.0 19.3 29.1 12.1 n 7 7 7 7 Rate of 41.984.9 inhibiting weight gain of colon (%)

In the above model, cell infiltration occurs with the colitis. Thus, thedegree of the cell infiltration is reflected in the colon weight and thetherapeutic effect on the inflammatory bowel disease can be evaluated bythe colon weight (see Ikenoue Y. et al., InternationalImmunopharmacology, 5: 993-1006, 2005).

<Inhibitory Effect on TNF-α Production> Extraction of Total RNA

The experimental diet obtained by mixing the amino acids shown in Table6 with the standard chow was given to the experiment mice, and afterthree weeks, the colon was removed. A colon sample was homogenized with500 μL of Isogen (Nippon Gene). The homogenate mixed with 100 μL ofchloroform was centrifuged at 15,000 rpm at 4° C. and an aqueous layerwas collected. An equivalent amount of 2-propanol was added thereto,which was then centrifuged at 15,000 rpm at 4° C. A resulting pellet waswashed with 70% (v/v) ethanol, dried in air and dissolved in watertreated with DEPC.

Synthesis of cDNA

All reagents used were from Invitrogen. 0.5 μg Of total RNA and 0.5 μgof oligo (dT) were dissolved in 12 μL of DEPC-treated water, heated at70° C. for 10 minutes, and cooled on ice for one minute. Subsequently, 4μL of 5×1st strand buffer, 1 μL of 10 mM dNTP mix and 2 μL of 100 mM DTTwere added thereto. The mixture was reacted at 42° C. for 5 minutes. 1μL (200 U) of SuperScriptII was added, and the mixture was reacted at42° C. for 50 minutes and 70° C. for 15 minutes.

Quantitative RT-PCR Using SYBR Green

A gene expression assay based on PCR was performed using SYBR Green PCRMaster Mix (Applied Biosystems) and ABI PRISM 7700 System. SYBR GreenPCR Master Mix comprises SYBR Green and heat resistant DNA polymerase,and detects fluorescence generated by specifically binding SYBR Green toa double strand DNA amplified by PCR using ABI PRISM 7700 System. Geneexpression amounts of the samples can be compared by comparing thenumbers of PCR cycles at which a significant fluorescence signal isdetected for the first time. The cDNA corresponding to 10 ng of totalRNA was used as a template of the quantitative PCR, and n=2 or more persample of measurements were performed. A forward primer and a reverseprimer at each 7.5 pmol of each gene were added to 7.5 μL of SYBR GreenPCR Master Mix (Applied Biosystems), and the total volume was made to be15 μL with water. The PCR was performed by 40 cycles of the reaction at95° C. for 10 seconds, 60° C. for 30 seconds and 72° C. for 30 secondsafter reacting at 95° C. for 10 minutes. The sequences of the primersused for the quantitative RT-PCR are as follows.

(SEQ ID NO: 1) Forward primer: CCACCACGCTCTTCTGTCTA (SEQ ID NO: 2)Reverse primer: AGGGTCTGGGCCATAGAACT

An inhibitory rate of TNF-α mRNA expression was calculated from thefollowing formula.

Inhibitory rate of TNF-α mRNA expression=[1−Relative amount of TNF-αmRNA expression in target group−Relative amount of TNF-α mRNA expressionin cell transfer group)/(Relative amount of TNF-α mRNA expression instandard chow group−Relative amount of TNF-α mRNA expression in celltransfer group)]×100(%)

Additionally, the amounts of mRNA for Reg3γ and Groα were also similarlymeasured. The following PCR primers were used.

Reg3 γ forward primer: AACAGAGGTGGATGGGAGTG (SEQ ID NO: 3) Reg3 γreverse primer: GGGTACCACAGTGATTGCCT (SEQ ID NO: 4) Gro αforward primer: GCTGGGATTCACCTCAAGAA (SEQ ID NO: 5) Gro αreverse primer: AAGGGAGCTTCAGGGTCAAG (SEQ ID NO: 6)

TABLE 6 Inhibitory rate of TNF-α mRNA expression Amino acid (%) 5% His54 5% Phe 84 30% Amino acid 49 mixture 2% Trp 33 5% Trp 49

TABLE 7 Amino acid Inhibitory rate of Reg3γ mRNA Expression (%) 5% His84

TABLE 8 Amino acid Inhibitory rate of Groα mRNA Expression (%) 5% His 70

1. An agent for treatment or prevention of an inflammatory bowel diseasewherein it contains one or more amino acids selected from the groupconsisting of lysine, histidine, phenylalanine, methionine, tryptophan,glutamine, glycine, cysteine, cystine and threonine, and the amino acidsare administered in an amount of 0.1 to 4000 mg/kg per day.
 2. The agentfor treatment or prevention of the inflammatory bowel disease of claim 1wherein it contains one or more amino acids selected from the groupconsisting of lysine, histidine, phenylalanine, tryptophan andglutamine.
 3. The agent for treatment or prevention of the inflammatorybowel disease of claim 1 wherein it contains tryptophan.
 4. The agentfor treatment or prevention of the inflammatory bowel disease of claim 3wherein it further contains one or more amino acids selected from thegroup consisting of lysine, histidine, phenylalanine and glutamine. 5.An agent for treatment or prevention of an inflammatory bowel diseasewherein it contains 0.5 to 2% by weight of tryptophan, 3 to 9% by weightof lysine, 1.5 to 5% by weight of histidine, 4 to 10% by weight ofphenylalanine and 9 to 23% by weight of glutamine, and the amino acidsare administered in an amount of 0.1 to 4000 mg/kg per day.
 6. The agentfor treatment or prevention of the inflammatory bowel disease of claim1, wherein the inflammatory bowel disease is Crohn's disease.
 7. Theagent for treatment or prevention of the inflammatory bowel disease ofclaim 1, wherein the inflammatory bowel disease is ulcerative colitis.8. An inhibitor of TNF-α production wherein it contains amino acidsselected from the group consisting of histidine, phenylalanine andtryptophan, and the amino acids are administered in an amount of 0.1 to4000 mg/kg per day.
 9. An inhibitor of TNF-α production wherein itcontains 0.5 to 2% by weight of tryptophan, 3 to 9% by weight of lysine,1.5 to 5% by weight of histidine, 4 to 10% by weight of phenylalanineand 9 to 23% by weight of glutamine, and the amino acids areadministered in an amount of 0.1 to 4000 mg/kg per day.